LOCUS       sequence-403638-        5611 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing human E2F2 protein with FLAG-tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5611)
  AUTHORS   Wang ME, Chen J, Lu Y, Bawcom AR, Wu J, Ou J, Asara JM, Armstrong 
            AJ, Wang Q, Li L, Wang Y, Huang J, Chen M
  TITLE     RB1-deficient prostate tumor growth and metastasis are vulnerable to
            ferroptosis induction via the E2F/ACSL4 axis.
  JOURNAL   J Clin Invest. 2023 Mar 16:e166647. doi: 10.1172/JCI166647.
  PUBMED    36928314
REFERENCE   2  (bases 1 to 5611)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Clin 
            Invest. 2023 Mar 16:e166647. doi: 10.1172/JCI166647."
COMMENT     Sequence Label: pCMV-Tag2B-E2F2
FEATURES             Location/Qualifiers
     source          1..5611
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        67..370
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        371..574
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     525..545
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        620..638
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      673..682
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             682..705
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2040..2056)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2041..2057)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(2112..2131)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2113..2131)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     polyA_signal    2405..2526
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2442..2461)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2496..2515
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2533..2988)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2670..2691)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2882..2901
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3015..3119
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3121..3478
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3121..3312
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3144..3164)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3329..3464
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3513..4307
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3513..4307
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3567..3586)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4177..4196
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4495..4514)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4539..4586
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4915..5503
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5404..5423
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atgcattagt tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga
       61 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg
      121 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg
      181 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
      241 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
      301 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
      361 tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
      421 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
      481 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
      541 gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta
      601 gcgattacgc caagctcgaa attaaccctc actaaaggga acaaaagctg gagctccacc
      661 gcggtggcgg ccgccaccat ggattacaag gatgacgacg ataagagccc gggcggatcc
      721 ctgcaagggc cccgggcctt ggcttcggcc gctgggcaga ccccgaaggt ggtgcccgcg
      781 atgagcccca cagagctgtg gccatccggc ctcagcagcc cccagctctg cccagctact
      841 gctacctact acacaccgct gtacccgcag acggcgcctc ccgcagcggc gccaggcacc
      901 tgcctcgacg ccactcccca cggacccgag ggccaagttg tgcgatgcct gccggcaggc
      961 cggctgccgg ccaaaaggaa gctggatctg gaggggattg ggaggcccgt cgtccctgag
     1021 ttcccaaccc ccaaggggaa gtgcatcaga gtggatggcc tccccagccc caaaaccccc
     1081 aaatcccccg gggagaagac tcggtatgac acttcgctgg ggctgctcac caagaagttc
     1141 atttacctcc tgagcgagtc agaggatggg gtcctggacc tgaactgggc cgctgaggtg
     1201 ctggacgtgc agaagcggcg catctatgac atcaccaacg tgctggaagg catccagctc
     1261 atccgcaaga aggccaagaa caacatccag tgggtaggca ggggaatgtt tgaagacccc
     1321 accagacctg ggaagcagca acagctgggg caggagctga aggagctgat gaacacggag
     1381 caggccttgg accatctcat ccagagctgc tctctgagct tcaagcacct gactgaggac
     1441 aaggccaaca agaggctggc ctatgtgact taccaggata tccgtgctgt tggcaacttt
     1501 aaggagcaga cagtgattgc cgtcaaggcc cctccgcaga cgagactgga agtgcccgac
     1561 aggactgagg acaacctgca gatatatctc aagagcaccc aagggcccat cgaagtctac
     1621 ctgtgcccag aggaggtgca ggagccggac agtccttccg aggagcctct cccctctacc
     1681 tccaccctct gccccagccc tgactctgcc cagcccagca gcagcaccga ccctagcatc
     1741 atggagccca cagcatcctc agtgccagca ccagcgccaa ccccccagca ggccccaccg
     1801 cctccatccc tggtcccctt ggaggctact gacagcctgc tggagctgcc gcacccactc
     1861 ctgcagcaga ctgaggacca gttcctgtcc ccgaccctgg cgtgcagctc ccctctgatc
     1921 agcttctccc catccttgga ccaggacgac tacctgtggg gcttggaggc gggtgagggc
     1981 atcagcgatc tcttcgactc ctacgacctt ggggacctgt tgattaattg aaagcttatc
     2041 gataccgtcg acctcgaggg ggggcccggt accttaatta attaaggtac caggtaagtg
     2101 tacccaattc gccctatagt gagtcgtatt acaattcact cgatcgccct tcccaacagt
     2161 tgcgcagcct gaatggcgaa tggagatcca atttttaagt gtataatgtg ttaaactact
     2221 gattctaatt gtttgtgtat tttagattca cagtcccaag gctcatttca ggcccctcag
     2281 tcctcacagt ctgttcatga tcataatcag ccataccaca tttgtagagg ttttacttgc
     2341 tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt
     2401 tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     2461 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     2521 atcttaacgc gtaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt
     2581 aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag
     2641 aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga
     2701 acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg
     2761 aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc
     2821 ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg
     2881 aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc
     2941 gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt
     3001 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     3061 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagaatc
     3121 ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     3181 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     3241 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     3301 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     3361 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
     3421 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaga
     3481 tcgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag
     3541 gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg
     3601 gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca
     3661 agaccgacct gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc
     3721 tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg
     3781 actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg
     3841 ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta
     3901 cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag
     3961 ccggtcttgt cgatcaggat gatctggacg aagaacatca ggggctcgcg ccagccgaac
     4021 tgttcgccag gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg acccatggcg
     4081 atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg
     4141 gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg
     4201 aagaacttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg
     4261 attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg
     4321 gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc
     4381 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     4441 ccagcgcggg gatctcatgc tggagttctt cgcccaccct agggggaggc taactgaaac
     4501 acggaaggag acaataccgg aaggaacccg cgctatgacg gcaataaaaa gacagaataa
     4561 aacgcacggt gttgggtcgt ttgttcataa acgcggggtt cggtcccagg gctggcactc
     4621 tgtcgatacc ccaccgagac cccattgggg ccaatacgcc cgcgtttctt ccttttcccc
     4681 accccacccc ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg gggcggcagg
     4741 ccctgccata gcctcaggtt actcatatat actttagatt gatttaaaac ttcattttta
     4801 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     4861 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     4921 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     4981 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     5041 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     5101 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     5161 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     5221 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     5281 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     5341 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     5401 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     5461 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     5521 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     5581 ccctgattct gtggataacc gtattaccgc c
//
